Metabolomics of Vitreous Humourfrom Retinoblastoma Patients

Importance of the Topic

Retinoblastoma is the most common pediatric ocular malignancy affecting children under five years old. The vitreous humour, situated between the lens and retina, can reflect tumor-related metabolic alterations due to its proximity to cancerous tissue.

Study Objectives and Overview

This study aimed to characterize differential metabolites in the vitreous humour of retinoblastoma patients compared to controls. A cohort of nine patient samples and two control samples was analyzed to link metabolic signatures with clinical risk categories.

Methodology

• Sample extraction using methanol:ethanol (1:1 v/v).
• LC-QTOF-MS in positive and negative ESI modes with C18 and HILIC columns.
• GC-QTOF-MS using derivatization and the Fiehn RTL method.
• Data processing with MassHunter Qualitative Analysis, Profinder, Agilent Unknown Analysis Software, and GeneSpring 13.1.
• Compound identification through METLIN, Fiehn RTL, and lipid annotation via SimLipid.

Used Instrumentation

• Agilent 6550 and 7200 Q-TOF mass spectrometers.
• Agilent 5975C single quadrupole mass spectrometer.
• ZORBAX RRHD SB-Aq and Poroshell 120 HILIC LC columns.
• DB-5MS GC column (PN: 122-5532G).
• Agilent MassHunter and GeneSpring software suites.

Key Results and Discussion

• Detection of over 1,000 molecular features spanning amino acids, carbohydrates, nucleobases, and lipids.
• High-risk patient samples showed distinct positive correlations and clustering apart from low-risk and control groups.
• Upregulation (up to fivefold) of phosphatidylcholines, ether-linked phosphatidylethanolamines, ceramides, sphingomyelins, and sphinganines in patient samples.
• Elevated levels of carnitines and free fatty acids suggest altered peroxisomal lipid metabolism.
• Volcano plot and hierarchical clustering identified approximately 350 significant metabolites (p ≤ 0.05, fold change ≥ 2.0).
• Pathway analysis revealed dysregulation in sphingolipid metabolism, glycolysis/gluconeogenesis, amino acid metabolism, ABC transporters, and HIF-1 signaling.

Benefits and Practical Applications

1. Identification of potential biomarkers for diagnosis and risk stratification in retinoblastoma.
2. Insights into tumor-associated lipid and energy metabolism.
3. Foundation for personalized chemotherapy and targeted therapies.

Future Trends and Opportunities

• Development of non-invasive sampling approaches, such as tear fluid analysis.
• Integration of multi-omics data for comprehensive tumor profiling.
• Creation of targeted metabolite panels for early detection and monitoring treatment response.
• Exploration of peroxisomal and lipid metabolism inhibitors as therapeutic interventions.

Conclusion

The integrated LC-QTOF-MS and GC-QTOF-MS metabolomic profiling of vitreous humour from retinoblastoma patients uncovered significant alterations in lipid and energy pathways, providing novel insights into tumor biology and potential diagnostic biomarkers. This multi-omics strategy enhances our understanding of disease mechanisms and supports future clinical translation.

References

• Gundimeda S., Lateef S., Guha N., Deepak S.A., Padmanaban A., Mallipatna A., Ghosh A. Metabolomics of Vitreous Humour from Retinoblastoma Patients. Metabolomics 2015; Poster 018.